Lulworthinone: In Vitro Mode of Action Investigation of an Antibacterial Dimeric Naphthopyrone Isolated from a Marine Fungus.

Treatment options for infections caused by antimicrobial-resistant bacteria are rendered ineffective, and drug alternatives are needed-either from new chemical classes or drugs with new modes of action. Historically, natural products have been important contributors to drug discovery. In a recent study, the dimeric naphthopyrone lulworthinone produced by an obligate marine fungus in the family Lulworthiaceae was discovered. The observed potent antibacterial activity against Gram-positive bacteria, including several clinical methicillin-resistant Staphylococcus aureus (MRSA) isolates, prompted this follow-up mode of action investigation. This paper aimed to characterize the antibacterial mode of action (MOA) of lulworthinone by combining in vitro assays, NMR experiments and microscopy. The results point to a MOA targeting the bacterial membrane, leading to improper cell division. Treatment with lulworthinone induced an upregulation of genes responding to cell envelope stress in Bacillus subtilis. Analysis of the membrane integrity and membrane potential indicated that lulworthinone targets the bacterial membrane without destroying it. This was supported by NMR experiments using artificial lipid bilayers. Fluorescence microscopy revealed that lulworthinone affects cell morphology and impedes the localization of the cell division protein FtsZ. Surface plasmon resonance and dynamic light scattering assays showed that this activity is linked with the compound's ability to form colloidal aggregates. Antibacterial agents acting at cell membranes are of special interest, as the development of bacterial resistance to such compounds is deemed more difficult to occur.

Microemulsions and nanoemulsions modified with cationic surfactants for improving the solubility and therapeutic efficacy of loaded drug indomethacin.

In this work, a noncovalent strategy was successfully used to modify colloidal stability andin vitroandin vivoefficacy of two amphiphilic formulations of the anti-inflammatory drug indomethacin. Namely, nanoemulsions and microemulsions based on oleic acid and nonionic surfactants have been produced and compared. The influence of cationic surfactants cetyltrimethylammonium bromide and its carbamate bearing analogue on the size characteristics, stability and ability to provide prolonged action of loaded drug indomethacin has been evaluated. Adding the positively charged molecules in the surface layer of nanoemulsions and microemulsions has shown the stability increase along with maintaining the size characteristics and homogeneity in time. Moreover, the carbamate modified analogue demonstrated beneficial behavior. Indomethacin loaded in microemulsions and nanoemulsions showed prolonged-release (10%-15% release for 5 h) compared to a free drug (complete release for 5 h). The rate of release of indomethacin from nanoemulsions was slightly higher than from microemulsions and insignificantly decreased with an increase in the concentration of the cationic surfactant. For carbamate surfactant nanocarrier loaded with fluorescence probe Nile Red, the ability to penetrate into the cell was supported by flow cytometry study and visualized by fluorescence microscopy.In vitrotests on anti-inflammatory activity of the systems demonstrated that the blood cell membrane stabilization increased in the case of modified microemulsion. The anti-inflammatory activity of the encapsulated drug was tested in rats using a carrageenan-induced edema model. Nanoemulsions without cationic surfactants appeared more efficient compared to microemulsions. Indomethacin emulsion formulations with carbamate surfactant added showed slower carrageenan-induced edema progression compared to unmodified compositions. Meanwhile, the edema completely disappeared upon treatment with emulsion loaded indomethacin after 4 h in the case of microemulsions versus 5 h in the case of nanoemulsions.

Utilizing Liposomal Quercetin and Gallic Acid in Localized Treatment of Vaginal Candida Infections.

Vulvovaginal candidiasis (VVC) is a widely spread fungal infection that causes itching, pain and inflammation at the vaginal site. Although common, currently available treatment suffers from limited efficacy and high recurrence. In addition, the growing problem of resistance to azole drugs used in current treatments emphasizes the need for superior treatment options. Antimicrobial polyphenols are an attractive approach offering multitargeting therapy. We aimed to develop novel liposomes for simultaneous delivery of two polyphenols (quercetin, Q, and gallic acid, GA) that, when released within the vaginal cavity, act in synergy to eradicate infection while alleviating the symptoms of VVC. Q was selected for its anti-itching and anti-inflammatory properties, while GA for its reported activity against Candida. Novel liposomes containing only Q (LP-Q), only GA (LP-GA) or both polyphenols (LP-Q+GA) were in the size range around 200 nm. Q was efficiently entrapped in both LP-Q and in LP-Q+GA (85%) while the entrapment of GA was higher in LP-Q+GA (30%) than in LP-GA (25%). Liposomes, especially LP-Q+GA, promoted sustained release of both polyphenols. Q and GA acted in synergy, increasing the antioxidant activities of a single polyphenol. Polyphenol-liposomes were not cytotoxic and displayed stronger anti-inflammatory effects than free polyphenols. Finally, LP-GA and LP-Q+GA considerably reduced C. albicans growth.

Amphipathic sulfonamidobenzamides mimicking small antimicrobial marine natural products; investigation of antibacterial and anti-biofilm activity against antibiotic resistant clinical isolates.

There is an urgent need for novel antimicrobial agents to address the threat of bacterial resistance to modern society. We have used a structural motif found in antimicrobial marine hit compounds as a basis for synthesizing a library of antimicrobial sulfonamidobenzamide lead compounds. Potent in vitro antimicrobial activity against clinically relevant bacterial strains was demonstrated for two compounds, G6 and J18, with minimal inhibitory concentrations (MIC) of 4-16 µg/ml against clinical methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE). The two compounds G6 and J18, together with several other compounds of this library, also caused ≥90% eradication of pre-established biofilm of methicillin-resistant S. epidermidis (MRSE) at 40 µg/ml. Using a luciferase assay, the mechanism of action of G6 was shown to resemble the biocide chlorhexidine by targeting the bacterial cell membrane.

Chitosan-Based Nanomedicine to Fight Genital Candida Infections: Chitosomes.

Vaginal infections are associated with high recurrence, which is often due to a lack of efficient treatment of complex vaginal infections comprised of several types of pathogens, especially fungi and bacteria. Chitosan, a mucoadhesive polymer with known antifungal effect, could offer a great improvement in vaginal therapy; the chitosan-based nanosystem could both provide antifungal effects and simultaneously deliver antibacterial drugs. We prepared chitosan-containing liposomes, chitosomes, where chitosan is both embedded in liposomes and surface-available as a coating layer. For antimicrobial activity, we entrapped metronidazole as a model drug. To prove that mucoadhesivness alone is not sufficient for successful delivery, we used Carbopol-containing liposomes as a control. All vesicles were characterized for their size, zeta potential, entrapment efficiency, and in vitro drug release. Chitosan-containing liposomes were able to assure the prolonged release of metronidazole. Their antifungal activity was evaluated in a C. albicans model; chitosan-containing liposomes exhibited a potent ability to inhibit the growth of C. albicans. The presence of chitosan was crucial for the system's antifungal activity. The antifungal efficacy of chitosomes combined with antibacterial potential of the entrapped metronidazole could offer improved efficacy in the treatment of mixed/complex vaginal infections.

Synthesis and antimicrobial activity of small cationic amphipathic aminobenzamide marine natural product mimics and evaluation of relevance against clinical isolates including ESBL-CARBA producing multi-resistant bacteria.

A library of small aminobenzamide derivatives was synthesised to explore a cationic amphipathic motif found in marine natural antimicrobials. The most potent compound E23 displayed minimal inhibitory concentrations (MICs) of 0.5-2µg/ml against several Gram-positive bacterial strains, including methicillin resistant Staphylococcus epidermidis (MRSE).E23 was also potent against 275 clinical isolates including Staphylococcus aureus, Enterococcus spp., Escherichia coli, Pseudomonas aeruginosa, and Klebsiella pneumoniae, as well as methicillin-resistant S. aureus (MRSA), vancomycin-resistant enterococci (VRE), and ESBL-CARBA producing multi-resistant Gram-negative bacteria. The study demonstrates how structural motifs found in marine natural antimicrobials can be a valuable source for making novel antimicrobial lead-compounds.